Species differ greatly in their rates of aging. Among mammalian species lifespan ranges from two to over sixty years. Long lifespan, and the slower rate of aging that leads to long lifespan, have evolved independently many times in diverse clades of mammals, to provide improved reproductive fitness in ecological niches that reward postponement of the diseases and disabilities of aging. Each evolutionary event that creates a long- lived species, or group of related species, presents new opportunities for learning about the biological factors that control aging rate. The proposed work seeks to better understand the biology behind interspecies lifespan differences. This work will be performed through a comparison of primary fibroblasts derived from a large number of species from a wide range of animal clades, where long lifespan has evolved independently. The decision to focus on fibroblasts does not require the (unlikely) assumption that age-related changes in fibroblast properties contribute in an important way to diseases that might occur in aged individuals. Instead, this approach is based on the assumption that evolutionary changes that produce slow aging might affect multiple cell types, including those which do contribute to long-lasting resistance to diseases and disabilities, and also those (like fibroblasts) which are easy to cultivate and expand under standardized conditions. The Miller lab has reported that fibroblasts from longer lived species are typically more resistant to oxidative stress than shorter lived species of the same clade. In initial work I have shown that cells from longer lived species of rodents, primates, birds and several other animal clades are less susceptible to protein damage following oxidative stress than cells from shorter lived species. In the work proposed in this training grant I plan to expand upon this finding and work out the cell biology behind these differences in resistance of the proteome to oxidative stress. This will be accomplished by comparisons of the levels and activity of specific components of the proteostasis system, known to be involved in oxidative stress resistance, using cell lines derived from approximately 100 long- and short-lived species from birds, rodents, primates, and laurasiatherian mammals. The work will involve measurements of the form and function of proteasome, mitochondrial Lon, antioxidant enzymes and molecular chaperones. A related Aim will focus on, and elucidate, our unexpected observation that cells from the longest lived species show a decline, below baseline levels, of protein carbonyl levels within one hour of oxidative stress, representing a fast-acting, inducible defense mechanism limited to cells from long-lived species.